ANALYSIS OF THE BINDING OF 3,3',5-TRIIODO-L-THYRONINE AND ITS ANALOGSTO MUTANT HUMAN BETA-1 THYROID-HORMONE RECEPTORS - A MODEL OF THE HORMONE-BINDING SITE

To understand the nature of the thyroid hormone binding site, we chara cterized the binding of 3,3',5-triiodo-L-thyronine (T3) and its analog ues to eight naturally occurring mutated human beta1 thyroid hormone r eceptors (h-TRbeta1). The mutant receptors were derived from patients with the syndrome of generalized thyroid hormone resistance, and each has a point mutation in the hormone binding domain (KT, R338W; TP, L45 0H; IR, D322H; NN, G347E; AH, P453H; OK, M442V; RL, F459C; and ED, A31 7T). Compared to the wild-type h-TRbeta1, binding of T3 was reduced by as much as 97% for the mutants. The order of binding affinity of wild -type h-TRbeta1 to the analogues is T3 > D-T3 > L-thyroxine > 3,5-diio do-L-thyronine > 3,3',5'-triiodo-L-thyronine. The mutant receptors sho wed essentially the same order of reduced affinities for the analogues , but the amounts of the reductions varied in each case. These results suggest specific local interactions (interplay) of analogues with the mutated residues in the receptors. On the basis of these data and a p utative structure of the hormone binding domain as an eight-stranded a lpha/beta barrel, we propose the location of the hormone in the bindin g site of h-TRbeta1. Ionic bonds anchor the hormone's alanine side cha in to loop 4 of the 8-fold alpha/beta barrel. The phenyl ring lies acr oss the amino-terminal face of the domain with the phenoxy ring pointi ng downward into the barrel interacting with beta-strand 8 on the oppo site side. Loops 1 and 7, which are located on the same face as the DN A binding domain, fold over the top of the barrel toward the bound hor mone. The T3-induced conformational changes observed by us and others may be partly mediated by loops 1 and 7. The present model should prov ide a basis for further studies to understand the T3-dependent transcr iptional activity of h-TRbeta1.